Apparatus for filtration-leukopheresis for separation and concentration of human granulocytes

ABSTRACT

Apparatus is provided for continuous withdrawal of blood from a human donor, separation preferably of granulocytes therefrom by extracorporeal circulation and filtration of the blood and return of leucocyte-poor whole blood to the donor, controls being provided for the flow of the blood within the apparatus so that the volume of the blood processed can be known and controlled at any particular time.

United States Patent 11 1 1111 3,802,32 Djerassi Apr. 9, 11974 APPARATUSFOR 3,228,397 1/1966 Moss 128/214 E FILTRATION LEUKOPHERESIS FOR3,656,478 4/1972 Swersey 128/214 E SEPARATION AND CONCENTRATION OF HUMANGRANULOCYTES Filed:

Inventor: Isaac Djerassi, 2034 Delancey Pl.,

Philadelphia, Pa. 19103 May 18, 1972 Appl. No: 254,609

References Cited UNITED STATES PATENTS Cordova 128/DIG. 3 Dyer 128/214 RGreenwalt et a1 128/214 R UX Judson et al. 128/214 R OTHER PUBLICATIONSIngram, Some Contributions of Leukocyte Balance Studies, U. ofRochester, 11/56, pp. 1-16.

Primary EraminerDalton L. Truluck Attorney, Agent, or Firn1Zachary T.Wobensmith,

[5 7] ABSTRACT Apparatus is provided for continuous withdrawal of bloodfrom a human donor, separation preferably of granulocytes therefrom byextracorporeal circulation and filtration of the blood and return ofleucocytepoor whole blood to the donor, controls being provided for theflow of the blood within the apparatus so that the volume of the bloodprocessed can be known and controlled at any particular time.

9 Claims, 3 Drawing Figures PATENTEDAPR 9 m4 3,802,432 saw 1 0f 2APPARATUS FOR FILTRATION-LEUKOPHERESIS FOR SEPARATION AND CONCENTRATIONOF HUMAN GRANULOCYTES BACKGROUND OF THE INVENTION 1. Field of theInvention This invention relates to apparatus forfiltrationleukopheresis for separation and concentration of largeamounts of normal human granulocytes, primarily for the purpose oftransfusion to individuals in need of such treatments.

2. Background of the Prior Art It has heretofore been proposed toextract blood from a human doner for use as whole blood, for separationof plasma and for separation of granulocytes or other blood components.

Blood removal and collection apparatus has taken various forms andreference may be had to the U.S. patents to Strumia, No. 2,845,929;Gewecke et al., No. 2,757,669; Welch, Jr., No. 2,982,286; Rieutord etal., No. 2,757,375; Portras, No. 2,784,932; Erikson, No. 2,597,715;Judson et al., No. 3,489,145.

The delivery of blood plasma and the like to a patient for transfusioncan be effected by gravity or by applied pressure as shown in the U.S.patent to Rundhaug, No. 2,842,123. The methods in current use forseparation and con centration of platelets and leukocytes fortransfusion rely on differential centrifugation. Note Judson et al.,U.S. Pat. No. 3,489,145.

The harvesting of normal granulocytes with special continuous flowcentrifugation (Freireich, EJ. et al., Separation and collection ofleukocytes. Cancer Res. :15 16-1520 1965) is difficult, costly and oflowefficiency.

it has heretofore been ascertained that granulocytes adhere to foreignsubstances such as siliconized glass wool and many of the newersynthetic plastics. A filter of the latter type and employing nylonfibers is available under the name Leuko Pak Leukocyte Filter fromFenwal Laboratories, Division of Travenol Laboratories, lnc., MortonGrove, Illinois. These filters have heretofore been used foradministration of leukocytepoor whole blood to multitransfused patientswith antileuko-cyte antibodies.

The adhering takes place when the medium carrying the granulocytes isheparinized blood and when the filter is perfused with more acid medium(example ACD or Citrate-Dextrose Plasma) the granulocytes are eluted andcan be recovered in the outflowing carrier fluid and then separatedtherefrom.

It has heretofore been undertaken to withdraw blood from a donor, in aplastic bag with a mixture of heparin, saline and sodium citrateselectively delivered to the plastic bag and then transferring the fluidthrough leukocyte filters and then advance the blood by manualmanipulation to another plastic bag for return through a tube to thedonor, a source of saline solution being connected to the return tube tokeep it open when not in use. The respective tubes were controlled byhemostats for manual regulation and the entire procedure was manual.

Very close supervision and control by trained personnelhas been requiredand the time for the filtrationleukopheresis has required about some 4hours or more with each individual donor.

Leukocyte transfusions using granulocytes obtained from patients withchronic myelogenous leukemia have been used to support patients withinfections who lack adequate numbers of granulocytes (Schwarzenberg, L.et al.: Study of factors'determining the'usefulness and complications ofleukocyte transfusions. Amer. J Med. 431206, 1967; and Morse, E.E. etal.: Effectiveness of granulocyte transfusion from donors with chronicmyelocytic leukemia to patients with leukopenia. Cl. Res. 9:32, 1961 Theuse of such transfusions is limited by the availability of donors withchronic myelogenous leukemia and the frequency of preexisting ordeveloping antileukocyte antibodies in the recipient leading to severeand often life-threatening transfusion reactions.

Normal leukocyte buffy coats have also been used for support of infectedleukopenic patients with inconclusive results. The transfusion of buffycoats from normal donors is handicapped by the need for each transfusionof large numbers of leukocyte concentrates (all ABO type specific) andespecially by their low content of granulocytes. The latter sediment oncentrifugation of the whole blood with the top layer of the red cellsand are not included in the buffy coat which consists mainly oflymphocytes and platelets.

Even if adequate yields of granulocytes are harvested from each unit ofwhole blood, large numbers of donors are needed for each singletransfusions thus greatly increasing the risk of sensitizing the patientto subsequent transfusions of granulocytes thus increasing the risk ofsevere reactions as well as of hepatitis and other infections.

Large amounts of granulocytes, all obtained from one donor as much aspossible compatible with the recipient, could however be givenrepeatedly to leukopenic patients in order to prevent or overcomealready existing infections. Obtaining normal granulocytes fortransfusion using continuous flow centrifuges is limited by the lowefficiency of separation by centrifugation (Freireich, E.J., et al.:Separation and collection of leukocytes. Cancer Res. 25:1516, 1965).Simple methods and suitable apparatus for repeated harvesting of largeamounts of granulocytes from single donors, applicable to standard bloodbanks and donor centers, are needed to use transfusions of granulocytesroutinely.

SUMMARY OF THE INVENTION In accordance with the present inventionapparatus is provided for harvesting granulocytes from a single donorand within a relatively short time period and which includes structurefor withdrawal of blood from the donor and transferred under pressure,the pressure application being effective to transfer the blood tofilters for retention of thegranulocytes by selectiveabsorption-elution. The filters are connected to leukocyte-poor wholeblood collecting receptacles from I which a connection is provided topumping apparatus blood to the donor which will be effective in itsaction, will protect the donor against excessive withdrawal of blood atany time, will insure effective filtration of the blood and its safereturn to the donor, which will reduce the burden upon the attendant andwhich will permit of relatively rapid cycling, thus allowing theprocessing of larger volumes of blood within the time available to thedonor.

It is a further object of the invention to provide apparatus of thecharacter aforesaid which will be reliable in its action, fool proof inregard to potential harm to the blood donor and which can be readilymade available for use.

Other objects and advantageous features of the invention will beapparent from the description and claims.

BRIEF DESCRIPTION OF THE DRAWINGS The nature and characteristic featuresof the invention will be more readily understood from the followingdescription taken in connection with the accompanying drawing formingpart hereof, in which:

FIG. 1 is a diagrammatic view of a preferred form of apparatus inaccordance with the invention;

FIG. 2 is a schematic diagram of the electrical circuitry employed; and

FIG. 3 is a view of a modified form of circuitry.

It should, of course, be understood that the description and drawingsherein are illustrative merely and that various modifications andchanges can be made in the structure disclosed without departing fromthe spirit of the invention.

DESCRIPTION OF THE PREFERRED EMBODIMENT Referring now more particularlyto FIG. 1 of the drawings, a fluid connection in the form of plastictubing is shown which has a needle (not shown) of well known type, andpreferably of gauge, on its free end for insertion into a vein in onearm A1 of the donor for withdrawing blood.

The fluid connection 10 preferably extends to the fluid inlet of anelectric motor driven pump 12. The outletor delivery connection of thepump 12 is connected by a fluid connection 14 to an elevated vessel 15suspended above the location at which the donor is situated. The pump 12can be of any desired type, rotary or tandem diaphragm or piston, but ispreferably of a variable speed type to provide the desired rate ofwithdrawal from the donor and delivery to the vessel 15.

The vessel 15 is connected by a fluid connection and branch pipes 21 toa plurality of leukocyte filters 22, four being preferred. The filters22 are connected by fluid connections 23 through infusion chambers 24which allow observation of the blood dripping or flowing out of theleukocyte filters after retention of the granulocytes. The filters 22can be of any suitable type which separate and retain granulocytes andpermit the passage of leukocyte-poor whole blood for return to thedonor, Leuko-pak Leukocyte Filters available from Fenwal Laboratories,Division of Travenol Laboratories, lnc., Morton Grove, Ill. having beenfound satisfactory.

The infusion chambers 24 are connected by interconnected branch tubes 25which is connected by a flexible tube 26 and flexible tubes 27 and 28 tocollection bags 29 and 30 which are resting on scales of the spring orlever type. The tubes 27 and 28 have solenoid controlled valves 32 and33 therein.

The collection bags 29 and 30 are carried on weigh- 5 ing scales 34 and35 having normally open switches 55 and 56 closed when a predeterminedweight is effective in either bag 29 or 30.

The collection bags 29 and 30 are connected by flexible pipes 35 and 36which have solenoid controlled valves 38 and 39 therein to a fluidconnection 40 which extends to the fluid inlet of an electric motordriven pump 42.

The pump 42 is preferably similar to the pump 12.

The delivery side of the pump 42 is connected by a 5 fluid connection 43to a vented receptacle 44 with a vent pipe 45 communicating with theatmosphere. The receptacle 44 has a fluid connection 46 extendingtherefrom which has a needle (not shown) of well known type andpreferably of 15 gauge on its free end for insertion into a vein in theother arm A2 of the donor for the return of granulocyte-poor whole bloodto the donor.

Referring now to FIG. 2, one form of electrical circuitry is shownsuitable for control of the system shown in FIG. 1.

A suitable source of electric energy such as l 15 volt 60 Hz alternatingcurrent is connected by a conductor 50 through a main power controlswitch SW 1 and conductor 51 and blood pump control switch SW2 conductor53, and contact arms 54 and 55 of a manually positioned blood pumpselector switch SW3 engageable with contacts 56 and 57 connected toconductors 58 and 59 to one terminal of each of the motor driven pumps12 and 42. The other terminals of the pumps 12 and 42 are connected toreturn conductor 60.

The contact arms 54 and 55 of SW3 can also be selectively connected tocontact 62 connected to conductor 58 and to contact 63 when operation ofpump 12 only is desired.

The contact arms 54 and 55 of the switch SW3 can also be selectivelyconnected to contact 64 and to contact 65 which is connected toconductor 59 when operation of pump 42 only is desired.

A spare blood pump can also be provided if desired and which can besubstituted by shifting of fluid inlet and delivery connections thereto.The pump 70 can be energized by normally open switch 71 connecting theconductor 53 through the pump 70 to the return line 60.

The conductors 51 and 60 are also preferably connected to the primary ofa step-down transformer T.

The secondary of the transformer T has one terminal thereof connected toa conductor 73 the other terminal being connected by a conductor 74through a normally open contact 75 of switch SW5 to one terminal ofsolenoid winding Kl thereof which has the other terminal connected toconductor 73. The secondary of the transformer T is also connected by aconductor 76 connected to the conductor 74 and through a normally opencontact 77 of switch SW6 to one terminal of winding of solenoid K2, theother terminal of which is connected to conductor 73.

The winding of solenoid Kl controls a contact arm 78 which in its downposition engages a contact 79 which is connected by a conductor 80 toone terminal of each of the solenoid valves 32 and 39.

The contact arm 78 in its up position engages a contact82 which isconnected by a conductor 84 to one terminal of the winding of solenoidK2, the other terminal of which is connected to conductor 73.

The winding of solenoid K2 controls a contact arm 85 which is connectedto conductor 74 and which in its up position engages a contact 86 whichis connected by a conductor 87 to conductor 74 and thence to oneterminal of the winding of the solenoid K1, the other terminal of whichis connected to conductor 73.

The winding of solenoid K2 controls the contact arm 85 to a downposition in engagement with a contact 88 which is connected by conductor89 to energize the windings of the solenoid valves 33 and 38.

The transformer T also has a conductor 90 which is connected byconductors 91, 92, 93, and 94 to the solenoid valves 32, 33, 38 and 39.

v A manually operable override SW7 with simultaneously operable contactarms 95 and 96 controls the independent energization of the solenoidvalves 32 and 33 for closing these valves.

A manually operable override switch SW8 with simultaneously operablecontact arms 97 and 98 controls the independent energization of thesolenoid valves 38 and 39 for closing these valves.

In normal operation with switches SW1 and SW2 closed, and with theswitch SW3 in normal operating position as shown in FIG. 2 the pumps 12and 42 will be operated. With the switch SW5 in closed position and theswitch SW6 in open position the solenoid Kl will be energized so thatsolenoid valves 32 and 39 will be closed for delivery of blood to thereceptacle 30, the valves 33 and 38 being open. As blood is delivered tothe receptacle 30 the switch SW6 will be closed to energize the solenoidK2. The flow of blood from the receptacle 29 and the closing of thecontact 77 by the weight of the blood in the receptacle 30 will effect areversal of the control of the valves 33 and 38 so that delivery to thereceptacle 44 through valve 39 will be effected with the valves 33 and38 closed. The alternating operation will be repeated as desired. a

The blood from the receptacle will pass by gravity through the filters24 and will have the granulocytes extracted therefrom for subsequentseparation and utilization.

The blood delivered by the pump 42 to the receptacle 44 is thenpreferably returned by gravity to the donor, any air in the blood beingseparated out in the receptacle 44. r

A modified form of circuitry is shown in FIG. 3 in which the scaleswitches includes a switch SWSa with normally open contact arm 75a andswitch SW6a with normally closed contact arm 77a to both of whichconductor 74 is connected. A single solenoid K is employed withconductor 90 from switch SWSa connected to one terminal thereof, theother terminal being connected to conductor 73. A conductor 91 extendsfrom switch SW6a to the normally closed contact arm 92 which whenengaged with contact 93 connected to conductor 90 for energizing thewinding of the solenoid K.

A normally down contact arm 94 when in engagement with a contact 95 isconnected by a conductor 96 to both the solenoid valve 32 and thesolenoid valve 39 6 for energizing the same, and when in an up positionin engagement with a contact 97 is connected by a conductor 98 to boththe solenoid valve 33 and the solenoid valve 38.

The alternating opening and closing of the valves controlling thedelivery to and discharge from the receptacles 29 and 30 is similar tothat previously described, initiated and continued by alternate closingof the scale switches SWSa and SW6a, these switches opening when theweight thereon is removed.

The processing of blood for separation and collection of granulocytes isgreatly facilitated.

I claim:

1. Apparatus for filtration-leukopheresis comprising a venous bloodsupply connection adapted to be connected to a donor,

a venous blood return connection adapted to be connected to the donor,

a leukocyte separating and retaining filter interposed in series withsaid connections and in fluid commu nication therewith for extractingleucocytes from blood passing therethrough,

a permanently vented receptacle in continuous communication with theatmosphere interposed in series between said filter and said returnconnection and in fluid communication therewith, and

power driven pump means upstream of the vented receptacle continuouslydelivering blood from said supply connection to and through said filterand to said vented receptacle.

2. Apparatus for filtration-leukopheresis comprising a blood supplyconnection adapted to be connected to a donor,

motor driven pump means for delivering blood from the supply connectionto a receptacle;

leukocyte separation filters to which blood from said receptacle isdelivered for circulation through said filters,

means for receiving blood from said filters including receptacles withvalved inlet and delivery connections,

weight responsive members associated with each of said second mentionedreceptacles,

members controlled by said weight responsive members for controlling theflow to and from said second mentioned receptacles, and

means for returning the blood from said delivery con nections to thedonor by increased gravity flow and venting of admixed air including ablood return connection adapted to be connected to the donor.

3. Apparatus for filtration-leukopheresis as defined in claim 2 in whichsaid last mentioned means includes a motor driven pump interposedbetween said blood return connection and said delivery connection forthe purpose to raise the blood to the level of the blood returnconnection and increase the flow by gravity back into the donor. 4.Apparatus for filtration-leukopheresis as defined in claim 2 in whichsaid blood return connection has air collecting and venting meansinterposed therein. 5. Apparatus for filtration-leukopheresis as definedin claim 2 in which said flow controlling members include solenoidcontrolled valves.

claim 5 in which independent control means is provided for the solenoidcontrolled valves for at least one of said filter connected receptacles.9. Apparatus for filtration-leukopheresis as defined in claim 2 in whichsaid leukocyte separation filters are positioned for flow of blood alsoby gravity through said filters.

1. Apparatus for filtration-leukopheresis comprising a venous bloodsupply connection adapted to be connected to a donor, a venous bloodreturn connection adapted to be connected to the donor, a leukocyteseparating and retaining filter interposed in series with saidconnections and in fluid communication therewith for extractingleucocytes from blood passing therethrough, a permanently ventedreceptacle in continuous communication with the atmosphere interposed inseries between said filter and said return connection and in fluidcommunication therewith, and power driven pump means upstream of thevented receptacle continuously delivering blood from said supplyconnection to and through said filter and to said vented receptacle. 2.Apparatus for filtration-leukopheresis comprising a blood supplyconnection adapted to be connected to a donor, motor driven pump meansfor delivering blood from the supply connection to a receptacle;leukocyte separation filters to which blood from said receptacle isdelivered for circulation through said filters, means for receivingblood from said filters including receptacles with valved inlet anddelivery connections, weight responsive members associated with each ofsaid second mentioned receptacles, members controlled by said weightresponsive members for controlling the flow to and from said secondmentioned receptacles, and means for returning the blood from saiddelivery connections to the donor by increased gravity flow and ventingof admixed air including a blood return connection adapted to beconnected to the donor.
 3. Apparatus for filtration-leukopheresis asdefined in claim 2 in which said last mentioned means includes a motordriven pump interposed between said blood return connection and saiddelivery connection for the purpose to raise the blood to the level ofthe blood return connection and increase the flow by gravity back intothe donor.
 4. ApParatus for filtration-leukopheresis as defined in claim2 in which said blood return connection has air collecting and ventingmeans interposed therein.
 5. Apparatus for filtration-leukopheresis asdefined in claim 2 in which said flow controlling members includesolenoid controlled valves.
 6. Apparatus for filtration-leukopheresis asdefined in claim 2 in which independent control means is provided forsaid first mentioned motor driven pump means.
 7. Apparatus forfiltration-leukopheresis as defined in claim 3 in which independentcontrol means is provided for said second mentioned motor driven pump.8. Apparatus for filtration-leukopheresis as defined in claim 5 in whichindependent control means is provided for the solenoid controlled valvesfor at least one of said filter connected receptacles.
 9. Apparatus forfiltration-leukopheresis as defined in claim 2 in which said leukocyteseparation filters are positioned for flow of blood also by gravitythrough said filters.